Pectin polymers are important constituents of plant primary cell walls. They are composed of chains of 1,4-linked .alpha.-D-galacturonic acid and methylated derivatives thereof. The use of pectin-degrading enzymes is important for the food industry, primarily in fruit and vegetable processing such as fruit juice production or wine making, where their ability to catalyse the degradation of the backbone of the pectin polymer is utilised.
An assortment of different pectin degrading enzymes is known to be present in various microorganisms such as Aspergillus niger. Of these, pectin methylesterase catalyses the removal of methanol from pectin, resulting in the formation of pectic acid (polygalacturonic acid). Pectate lyase cleaves glycosidic bonds in polygalacturonic acid by .beta.-elimination, pectin lyase cleaves the glycosidic bonds of highly methylated pectins by .beta.-elimination, and polygalacturonase hydrolyses the glycosidic linkages in the polygalacturonic acid chain.
More specifically, pectin lyase may be used, alone or in combination with one or more other pectin degrading enzymes, in the production of fruit juice, in particular citrus juice, for partial or complete degradation of the pulp present in the juice after pressing.
For many purposes, it would be desirable to provide each of the pectin degrading enzymes present in, for instance, commercial preparations containing a number of different pectin degrading enzymes (an example of such a preparation is Pectinex Ultra SP.RTM., prepared from Aspergillus aculeatus, available from Novo Nordisk A/S) in a form free from other components. In this way, it would be possible to produce enzyme preparations adapted to specific purposes, such preparations either containing a single pectin degrading enzyme or arbitrary combinations thereof. To serve this end, it is convenient to provide single-component pectin degrading enzymes by recombinant DNA techniques.
Plastow et al., Symbiosis 2, 1986, pp. 115-122, describe the cloning of four pectate lyase genes and one polygalacturonase gene from Erwinia in E. coli.
Cloning of a pectin lyase is described in EP 278 355 and EP 353 188.
M. Kusters-van Sommeren et al., Mol. Gen. Genet. 234, 1992, pp. 113-120, describe the nucleotide sequence of the Aspergillus niger pelB gene encoding pectin lyase B.
C. Gysler et al., Gene 89, 1990, pp. 101-108, describe the isolation and nucleotide sequence of the Aspergillus niger pelD gene encoding pectin lyase D.
A. Alana et al., FEBS Letters 280, 1991, pp. 335-340, describe the purification and some characteristics of a pectin lyase from Penicillium italicum.